Toxoplasma gondii is an important opportunistic infection in AIDS patients and other immune- compromised individuals as well as fetuses. The parasite causes a broad spectrum of disorders in a variety of tissues including the brain and retina. The parasite has a complex life cycle during which it encounters diverse O2 environments. These include tissues with low O2 tensions such as the intestine, which is the first tissue that comes in contact with the parasite, and peripheral tissues such as regions of the brain that are distant from the vasculature. Adapting to changes in O2 availability is a critical cellular process for all respiring cells and organisms. How Toxoplasma senses O2 is unknown but our published and additional preliminary data indicate that this is critical for parasite growt. Cytoplasmic prolyl 4-hydroxylases (PHDs) are key cellular O2 sensors in metazoans and other protists that function by catalyzing the transfer of one atom from O2 to a protein-linked proline residue and the second one to ?-ketoglutarate yielding succinate, CO2 and a hydroxylated proline residue. We have identified two PHDs in Toxoplasma and our data suggests that one of these TgPhyB is an essential enzyme that can hydroxylate SKP1, which is a subunit of the E3SCF ubiquitin ligase complex. The goal of this proposal is to define the essentiality of TgPhyB, determine its substrate specificity, and identify TgPhyB inhibitors. Successful completion of these aims will unveil a novel signaling mechanism in Toxoplasma and establish how and why Toxoplasma O2 sensing is important for growth. Most importantly, this work could potentially reveal a new drug target to treat this critical opportunistic infection of AIDS patients.